In addition to our work in pancreas, identification of the sites of CRHSP-24 phosphorylation provides insight to the role of calcineurin and other phosphatases in cell culture based assays for growth, differentiation, adhesion and cell migration. Global phosphoproteomic approaches have revealed regulated phosphorylation of CRHSP-24 serines 30, 32, and 53 in growing and differentiating C2C12 muscle
apexbio and identified a unique phosphorylation site at Ser2[18] not seen in pancreas (this study) or HEK293 cells [12]. Although CRHSP-24 was not a focus of the work, calcineurin activity is known to be important for normal muscle development and in limiting the pathogenesis of dystrophic myofibers [19]. By contrast, CRHSP-24 Ser42 appears constitutively phosphorylated in C2C12 cells [18] and phosphorylation at this site has been observed during integrin-mediated adhesion and spreading of HeLa
cells [20]. Interestingly, the Drosophila melanogaster ortholog of CRHSP-24, CG9705 (Fig. 4), was found to bind the corresponding fly proteins for calmodulin and fermitin in global yeast two-hybrid screens of protein interactions [21]. A potential interaction with calmodulin may target CRHSP-24 for subsequent Ca2+-dependent dephosphorylation by calcineurin, whereas, fermitin homologues of human and flies are found as components of integrin-containing cell adhesion structures involved in muscle assembly and maintenance [22]. Interestingly, all of the serine phosphorylation sites identified in this study are conserved in fly CG9705 (Fig. 4), thereby providing the basis to study the role of CRHSP-24 phosphorylation in regulating calmodulin and fermitin interaction in this organism.
